Characterization of G protein-coupled receptors displayed on bio-nano magnetic particles.

Accession number;04A0258601
Title;Characterization of G protein-coupled receptors displayed on bio-nano magnetic particles.
Author; YOSHINO TOMOKO (Tokyo Univ. Agric. and Technol., Fac. Technol., JPN) TAKEYAMA HARUKO (Tokyo Univ. Agric. and Technol., Fac. Technol., JPN) TANAKA TSUYOSHI (Tokyo Univ. Agric. and Technol., Fac. Technol., JPN) MATSUNAGA TADASHI (Tokyo Univ. Agric. and Technol., Fac. Technol., JPN)
Journal Title;Abstracts. Symposium on Biofunctional Chemistry
Journal Code:L0836A
ISSN:
VOL.18th;NO.;PAGE.36-37(2003)
Figure&Table&Reference;
Pub. Country;Japan
Language;Japanese
Abstract;G protein-coupled receptors (GPCRs) play central roles in a wide range of biological processes, and are prime targets for drug discovery. GPCRs have large hydrophobic domains, thus the process for purification from the cell membrane are frequently time-consuming and typically results in loss of native conformation. In this study we assembled GPCRs on nano-sized bacterial magnetic particle (BMPs) by genetic engineering using Magnetospirillum magneticum AMB-1. Mms16, which is abundantly produced on the BMP membrane, was used as an anchor molecule which helps the functional protein to be localized on BMPs. The GPCR genes with or without Mms16 were cloned downstream of the mms16 promoter in an expression vector. Constructed vectors were introduced into wild-type AMB-1. Only transformant harboring Mms16 gene grew under antibiotics. Cytoplasmic fraction, cell membrane fraction, and BMP fraction were separated, and expression of GPCRs in cell membrane and BMP fractions was confirmed by ELISA using anti-GPCR antibody. Saturation binding analysis was used to assess the affinity of ligand to GPCR-BMP complexes. GPCR on BMPs showed a single affinity for its ligand. This system makes possible the convenient acquisition of the native conformation of GPCRs without for detergent solubilization, purification and reconstitution after cell disruption. (author abst.)
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